The present invention generally relates to the regulation of plant growth and to methods of inhibiting ethylene responses in plants by application of cyclopropene, cyclopentadiene, diazocyclopentadiene or their derivatives, in particular methylcyclopropene. The present invention specifically relates to methods of synthesis and molecular encapsulation agent complexes, in addition to storage, transport and application of these gases that inhibit ethylene responses in plants.
Plant growth responses are affected by both internal and external factors. Internal control of plant processes are under the influence of genetic expression of the biological clocks of the plant. These processes influence both the extent and timing of growth processes. Such responses are mediated by signals of various types which are transmitted within and between cells. Intracellular communication in plants typically occurs via hormones (or chemical messengers) as well as other less understood processes.
Because communications in a plant are typically mediated by plant hormones, both the presence and levels of such hormones are important to specific plant cell reactions. The plant hormone that is most relevant to the present invention is ethylene, which has the capacity to affect many important aspects of plant growth, development and senescence. The most important effects of ethylene include processes normally associated with senescence, particularly fruit ripening, flower fading and leaf abscission.
It is well known that ethylene can cause the premature death of plants including flowers, leaves, fruits and vegetables. It can also promote leaf yellowing and stunted growth as well as premature fruit, flower and leaf drop.
Because of these ethylene-induced problems, very active and intense research presently concerns the investigation of ways to prevent or reduce the deleterious effects of ethylene on plants.
One major type of treatment used to mitigate the effects of ethylene employs ethylene synthesis inhibitors. These ethylene synthesis inhibitors reduce the quantity of ethylene that a plant can produce. Specifically, these ethylene synthesis inhibitors inhibit pyridoxal phosphate-mediated reactions and thereby prevent the transformation of S-adenosynimethione to 1-amino cyclopropane-1-carboxylic acid, the precursor to ethylene. Staby et al. ("Efficacies of Commercial Anti-ethylene Products for Fresh Cut Flowers", Hort Technology, pp. 199-202, 1993) discuss the limitations of these ethylene synthesis inhibitors. Because ethylene synthesis inhibitors only inhibit a treated plant's production of ethylene, they do not suppress the negative effects of ethylene from environmental sources. These environment sources of ethylene exist because ethylene is also produced by other crops, truck exhaust, ethylene gashing units and other sources, all of which can affect a plant during production, shipment, distribution and end use. Because of this, ethylene synthesis inhibitors are less effective than products that thwart a plant's ethylene responses. For a discussion of the ethylene response in plants, see U.S. Pat. No. 3,879,188.
The other major type of treatment used to mitigate the effects of ethylene employs blocking the receptor site that signals ethylene action. One of the best known compounds for inhibiting the ethylene response in plants, as well as preventing the deleterious effects from environmental sources of ethylene, is silver thiosulfate ("STS"). An example of a commercial STS product is SILFLOR solution, available from Floralife, Inc., Burr Ridge, Ill. STS is very effective in inhibiting the ethylene response in plants and has been used because it moves easily in the plant and is not toxic to plants in its effective concentration range. STS can be used by growers, retailers and wholesalers as a liquid that is absorbed into the stems of the flowers. While STS is highly effective, it has a serious waste disposal problem. It is illegal to dispose of the silver component of STS by conventional means, such as by using a laboratory sink, without first pretreating the STS to remove the silver. It is also illegal to spray STS on potted plants. Consequently because of this disposal problem which is typically ignored by growers, STS is now almost exclusively utilized only by growers. Therefore, there is a great desire among postharvest physiologists to find alternatives to STS. To the knowledge of the present inventors, the only commercially acceptable replacements for STS are cyclopropene, cyclopentadiene, diazocyclopentadiene and their derivatives.
Many compounds such as carbon dioxide which block the action of ethylene diffuse from the ethylene receptor or binding site over a period of a few hours. Sisler & Wood, Plant Growth Reg. 7, 181-191, 1988. While these compounds may be used to inhibit the action of ethylene, their effect is reversible and therefore they must be exposed to the plant in a continuous manner if the ethylene inhibition effect is to last for more than a few hours. Therefore, an effective agent for inhibiting the ethylene response in plants should provide an irreversible blocking of the ethylene binding sites and thereby allow treatments to be of short duration.
An example of an irreversible ethylene inhibiting agent is disclosed in U.S. Pat. No. 5,100,462. However, the diazocyclopentadiene described in that patent is unstable and has a strong odor. Sisler et al., Plant Growth Reg. 9, 157-164, 1990, showed in a preliminary study that cyclopentadiene was an effective blocking agent for ethylene binding. However, the cyclopentadiene described in that reference is also unstable and has a strong odor.
U.S. Pat. No. 5,518,988 discloses the use of cyclopropene and its derivatives, including methylcyclopropene, as effective blocking agents for ethylene binding. Although the compounds in this patent do not suffer from the odor problems of diazocyclopentadiene and cyclopentadiene, because they contain a carbene group, they are relatively unstable due to their potential for undergoing oxidation and other reactions. Therefore, a problem of stability of these gases, as well as the explosive hazards these gases present when compressed, exist. To solve these problems, the present inventors have developed a method of incorporating these gaseous compounds, which inhibit the ethylene response in plants, in a molecular encapsulation agent complex in order to stabilize their reactivity and thereby provide a convenient and safe means of storing, transporting and applying or delivering the active compounds to plants. The application or delivery methods of these active compounds can be accomplished by simply adding water to the molecular encapsulation agent complex.
In trying to implement the teaching of U.S. Pat. No. 5,518,988, the problems associated with the stability of the gases and the potential explosive hazard of using compressed gases limit their use and therefore their effectiveness. To solve those problems, the present inventors developed a molecular encapsulation agent complex that stabilizes the reactivity of these gases and thereby provides a convenient and safe means of storing, transporting and applying or delivering these gases to plants.
This approach is an important advance over the art as it allows for the convenient and safe storage, transport and use of gases that are otherwise difficult to store, ship and dispense. The present invention will now allow for the safe, convenient and consistent use of these gases in the field by the grower, in addition to their use in distribution and in the retail marketplace. In fact, a complex of methylcyclopropene and the molecular encapsulating agent cycloclextrin allows for a product having a shelf life of greater than one year.
Another feature of the molecular encapsulation agents of the present invention is that once they trap the gaseous active agent in the complex, the complex (and hence the gaseous active agent) does not exhibit a very high vapor pressure and is therefore protected from oxidation and other chemical degradation reactions. A gaseous active compound such as cyclopropene or derivatives thereof is held in a caged molecule whereby the vapor pressure of the solid is very low due to the weak atomic forces (van de Waals and hydrogen binding). The binding of these gaseous active compounds with these molecular encapsulation agents holds the active compound until ready for use.
The present invention also prolongs the life of plants by providing an effective and proper dose of the encapsulated active compound capable of inhibiting the ethylene response, which is subsequently desorbed into a gas form for administration to the plant. The invention further embodies the release of the desired active compound from the complex by dissolving the complex in a suitable solvent in order to release the gaseous active compound, thereby serving as an improved gaseous plant treatment.
A major advantage of the present invention is that it provides an effective, user-friendly product for non-technical customers, florists and wholesalers. In addition, the molecular encapsulation agent complex acts as a controlled release agent for treatment with such active gaseous compounds as cyclopropene and methylcyclopropene. As a result, the present invention promotes less human exposure to the target compound than other means of application. Additionally, the user has more control over the application of the gaseous active compound because the active gaseous compound is slowly released from the complex in the presence of a suitable solvent.
Another advantage of the present invention is the amount of selective inclusion of the gaseous active compounds such as cyclopropene and methylcyclopropene into the molecular encapsulation agent. Using the teachings of the present invention, significant quantities of methylcyclopropene and other active compounds can now be encapsulated into a molecular encapsulation agent such as cyclodextrin, far exceeding the normal expected amount usually found with other solids.
A still further advantage of the present invention over the use of compressed concentrated gases is the elimination of the need for gas tanks, regulators, and OSHA compliance for pressurized gas tanks. This results in a substantial cost savings for the manufacturer as well as the customer. In addition, it eliminates the explosive and flammable potential associated with the use of gas tanks holding a highly reactive organic molecule. Moreover, the present invention eliminates the self polymerization and decomposition of gases that occur with compressed gases or liquids containing them.
Another advantage of the present invention over other inert solid carrier systems proposed for use in applying cyclopropene, such as dust, talc, silica and flour, is that it provides a product containing the active gaseous compound with increased stability. For example, the molecular encapsulation agent cyclodextrin protects the active cyclopropene or methylcyclopropene molecule from external conditions, such as ultraviolet degradation, which are problematic in photosensitive compounds such as these.
A still further advantage of the present invention is that the molecular encapsulation agent complex results in more effective use of the active gaseous compound. For example, a reduced quantity of cyclopropene can be utilized to obtain an effective treatment compared with the use of prior proposed cyclopropene solid carriers or compressed gases. This results in less waste and less packaging needed for the commercial product.
In another embodiment, this invention relates to the synthesis of cyclopropene and its derivatives including methylcyclopropene by methods that lower the incidence of impurities, such as hazardous reaction products and by-products, that interfere with the ethylene binding effectiveness of cyclopropene and its derivatives. These reaction product impurities include compounds that bind tightly but reversibly to the ethylene receptor site and inhibit the irreversible binding of cyclopropene and its derivatives, especially methylcyclopropene. The synthesis of these cyclopropene and derivative compounds is important because if irreversible binding to the receptor site does not take place during plant treatment, the plant will not be protected against the effects of ethylene.
The prior art syntheses of methylcyclopropene has created problems when the methylcyclopropene was used for inhibiting the ethylene response in plants. While it is well documented in U.S. Pat. No. 5,518,988 that methylcyclopropene and other similar compounds are active against ethylene, it has been discovered that not all methods of synthesis are as effective or preferable as the presently claimed synthesis method.
First, it is necessary to avoid producing during synthesis products (or impurities) that reversibly bind to the same ethylene receptor site as the intended active compound. Because these impurities do not irreversibly bind in a manner consistent with the inactivation of the receptor site without phytotoxicity, the effectiveness of using such a reaction product mixture without further processing is reduced. The specific impurities that must be avoided in the synthesis in order to obtain optimal performance of the reaction mixture include methylenecyclopropane, methylcyclopropanes and butanes.
The present inventors have discovered that of all the Lewis bases used for the production of methylcyclopropene, sodium amide and lithium diisopropylamide are most preferred. Synthesis using various metal hydrides and hydroxides were found to produce high levels of other reaction products that lowered the performance of the methylcyclopropene for plant uses. For example, using butynes, 3-hydroxy-2-methylpropenes and other similar starting materials generally yields an impure reaction product that is not appropriate for use in the treatment of plants.
Additional features and advantages of the present invention are described in, and will be apparent from, the detailed description and examples provided.